Flow cytometry photomultiplier tubes

Fluorescence excitation with a laser microbeam allows for a smaller region to be illuminated. Monitoring fluorescence with a sensitive photomultiplier tube also permits the use of lower intensities of irradiation for shorter periods of time. Therefore, unwanted photobleaching can be significantly reduced. If the spot size is adjusted to illuminate an entire cell, information analogous to spectrofluorometry or flow cytometry can be obtained on an individual cell basis with a high degree of temporal resolution. If the spot size is smaller than the cell, similar information can be obtained from a particular location within the cell. 

Figure 1. Schematic diagram of flow cytometry measurement of the frequency function, f, for single-cell protein content, p, in a cell population. After cellular protein is stained with a specific fluorochrome, the dilute cell suspension is analyzed in the flow cytometer. Laser radiation excites stain fluorescence in a single cell that is measured by a photomultiplier tube. The resulting voltage pulses are sorted and stored to obtain the frequency function.

Flow cytometry is used to separate populations of cells in a mixture from one another by means of fluorescently labeled antibodies and DNA-specific dyes. Antibodies used are usually against the molecules expressed on the cell surface, such as cell-surface receptors. The labeled cells are diluted in a solution and passed through a nozzle in a single-cell stream while being illuminated by a laser beam. Fluorescence is detected by photomultiplier tubes (PMTs) with optical filters to determine the emission wavelength, which in turn helps to identify the surface receptors (Figure 8.9). Fluorescence data are used to measure the relative proportions of various cell types in the mixture, and to derive the diagnostic information [22]. 

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